The use of sep-pak[superscript r] cartridges in aldosterone determinations using high-pressure liquid chromatography

Abstract

Aldosterone is a steroid which is essential to life. Because of its clinical significance, it is important that an accurate method be utilized when assaying its level in urine. Most clinical laboratories have use RIA procedures when analyzing urines for this steroid. One of the non-isotope procedures investigated for the determination of aldosterone is high-pressure liquid chromatography. Prior to the actual quantification of this steroid by HPLC, it must be extracted from the urine and hydrolyzed. This has usually been accomplished by tedious, time-consuming methods.

The purpose of this research was to devise a procedure which would quickly and completely extract aldosterone from urine specimens, while at the same time, eliminating interferring compounds. this study utilized Waters Associates, Inc. Sep-Pak silica and C18 cartridges. By utilizing these cartridges in sequence, undesirable components, such as urinary, pigments can be removed. Recovery of aldosterone standard added to unhydrolyzed random urine was 53-73%. Operating parameters include a reverse-phase C18 column and aqueous acetonitrile as the mobile phase.

Difficulties arose after acid hydrolysis of the urine sample was completed. There appears to be some type of problem occurring which prevents the extraction of aldosterone by theses Sep-Pak cartridges under the particular operating parameters used. Suggestions for dealing with this problem are discussed.